The efficacy and safety of oral microecological agents as add‐on therapy for atopic dermatitis: A systematic review and meta‐analysis of randomized clinical trials

Abstract Background Atopic dermatitis (AD) is a common skin disease that is hard to completely cure in a short time. Guidelines recommend the use of topical corticosteroids (TCS) as first‐line anti‐inflammatory therapy for AD, but long‐term use has significant side effects. Microecological agents (MA), including probiotics, prebiotics and synbiotics, have been widely reported as a potential adjunctive therapy of AD, but whether MA can contribute to AD treatment is currently controversial. Therefore, we conducted a systematic review and meta‐analysis to investigate whether MA as an add‐on therapy for AD has synergistic and attenuated effects and to further understand the role of MA in clinical interventions for AD. Methods We systematically searched Medline, Embase, Web of Science, Cochrane Library and PsycINFO databases up to Apr 11, 2023, and bibliographies were also manually searched, for potentially relevant studies regarding MA as additional therapy of AD. The Cochrane Risk of Bias Tool for assessing risk of bias was used to assess the quality of randomized controlled trials (RCTs). Two reviewers screened studies, extracted data, and evaluated the risk of bias independently. The primary outcomes (SCORAD scores and the number of adverse events) and the secondary outcomes (pruritus scores, the quality of life and the frequency of TCS) were extracted from each article. The data were combined and analyzed to quantify the safety and efficacy of the treatment. R (V4.4.3) software was used for data synthesis. The certainty of the evidence was evaluated with the Grade of Recommendation, Assessment, Development and Evaluation (GRADE) system. We also performed a trial sequential analysis to assess the reliability of the evidence. Results A total of 21 studies, including 1230 individuals, were identified, 20 of which met the eligibility criteria for the meta‐analysis. Our pooled meta‐analyses showed that compared with controls, oral MA as an add‐on therapy was associated with significantly lower SCORAD scores (MD = −5.30, 95% CI −8.50, −1.55, p < 0.01, I 2 = 81%). However, adverse events, pruritus scores, quality of life, and frequency of TCS use showed no significant difference in this meta‐analysis study (p > 0.05). Conclusions This meta‐analysis showed that MA plus TCS could be an effective and safe treatment for patients with AD to relieve relevant symptoms, which might be used as an add‐on therapy in the treatment of AD. However, due to the limited number of studies, results should be interpreted with caution. Further studies with a larger sample size are needed to explore the optimal protocol of MA plus TCS.


| INTRODUCTION
Atopic dermatitis (AD), characterized by intense itching, dry skin and redness, is a chronic relapsing inflammatory skin disease.It is one of the most common skin diseases in dermatological practice. 1The global prevalence of AD ranged from 15% to 20% in children and up to 10% in adults. 2 AD had the highest disability adjusted life year (DALY) burden of all skin diseases and ranked the 15th among all nonfatal diseases globally. 3In addition, AD is commonly associated with sleep disturbances, negative emotions, and decreased productivity, which seriously affects the patients' quality of life. 4AD has become one of the most intractable public health issues worldwide.
The basic management of AD involves emollient therapy, topical therapy, avoiding specific and non-specific triggers. 5The ETFAD/ EADV Eczema task force 6 and the European guidelines 5 recommended topical corticosteroids (TCS) as the first-line antiinflammatory treatment for AD.Although TCS has been the mainstay of AD treatment, while its long-term and frequent use is accompanied by several adverse effects, for example, skin atrophy, stria, purpura, hypothalamic pituitary axis suppression and growth suppression. 7Inappropriate TCS use may induce topical steroid withdrawal syndrome. 8Furthermore, corticophobia was also common in patients with AD who mistakenly believed that the side effects of TCS outweighed its therapeutic benefits. 9This would decrease treatment adherence and limit the possibilities for better control of the symptoms.To avoid steroid-related side-effects and reduce the lengthy use, application of TCS in combination with other treatment modalities was recommended 6 by the guidelines.
Microecological agents (MA) including probiotics, prebiotics and synbiotics are able to modulate gut microbiota, improve gut barrier function and relieve symptoms of skin diseases. 10Results of clinical studies 11,12 showed that MA supplementation could improve symptomatology, clinical severity of AD and quality of life.
Moreover, researchers suggested that MA had a steroid sparing effect, could be used as an add-on therapy to TCS. 13,14 Although studies have been conducted on the effect of MA as an add-on therapy for AD, these results are conflicting. 12,15Besides, MA related adverse events included abdominal pain, diarrhea, etc. 16 Therefore, we raised the following questions as an add-on therapy: (1) whether MA plus TCS is more effective than TCS alone?(2)   whether MA is able to reduce the accompanying side effects of TCS or the dependence on TCS? (3) what are the influencing factors on the effect of MA?

| MATERIALS AND METHODS
We conducted this systematic review and meta-analysis according to the A Measurement Tool to Assess Systematic Reviews 2 (AMSTAR 2.0) 17 and reported conforming to the Preferred Reporting Items for Systematic Evaluations and Meta-Analyses (PRISMA) guidelines. 18e protocol for this systematic review and meta-analysis was registered in the International Prospective Register of Systematic Reviews (PROSPERO) as CRD42023426811(https://www.crd.york.ac.uk/prospero/#myprospero).

| Data sources
We systematically retrieved the randomized controlled trials (RCTs) of MA as an add-on therapy to TCS for AD from Medline, Embase, Web of Science, Cochrane library, and PsycINFO from their inception to Apr 11, 2023, using the following terms: "Atopic Dermatitis", "probiotics", "prebiotics" and "synbiotics".Furthermore, gray literature, the references of identified RCTs, relevant reviews and clinical registration websites (ClinicalTrials.gov)were also searched to identify additional RCTs.We consulted experts for possible eligible studies.The entire search strategies were constructed by an experience reviewer (JL) and the detailed search strategies are provided in Appendix 1.

| Eligibility criteria
To be included for systematic review: (1) Participants were diagnosed as AD based on the Hanifin and Rajka criteria, 19 with no limitation of age or gender.(2) Oral probiotics or prebiotics or synbiotics were used as an add-on therapy to TCS. (3) The comparison included TCS alone, or TCS in combination with placebo.(4) The primary outcomes were SCORAD scores and adverse events.Secondary outcomes were pruritus scores, the frequency of TCS and quality of life.Among which, SCORAD scores or adverse events were obligatory.(5)   Double-blind RCTs.(6) Full texts were available.(7) Studies were published in English.
To be excluded for systematic review: (1) data were inaccessible, (2) non-double-blinded cross-over RCTs, reviews, duplicated publications, commentaries, case reports, case series, observational studies, comments, or in vitro studies.

| Studies selection
Endnote X9 was used to manage the retrieved records.After removing duplicates, two independent reviewers (HYQ and PWX) screened titles and abstracts based on inclusion and exclusion criteria.Then, full texts were reviewed to determine eligible studies.
After that, the included studies were cross-examined.Any disagreement was resolved by consulting a third reviewer (JL).

| Data collection and extraction
Two reviewers (DQ and PWX) independently extracted data using a predefined data extraction form.The following data were extracted: With regards to missing data, the corresponding authors were contacted via email.If the data were not displayed by mean and standard deviation, the formula recommended by the Cochrane handbook 20,21 was used to convert the data.In the case of the data presented in graphs, the semi-automated extraction tool WebPlotDigitizer (https://automeris.io/WebPlotDigitizer/,Version 4.3) was utilized to extract data. 22,23

| Risk of bias assessment
The revised Cochrane risk-of-bias tool for randomized trials (ROB 2.0) was used to evaluate the risk of bias from five domains: randomization process, deviation from the intended intervention, missing outcome data, measurement of the outcome, and selection of the reported results. 24Each domain was judged as "low risk," "some concerns," or "high risk".Two reviewers (PWX and HLL) independently assessed the risk of bias and then cross-checked.After crossexamination, disagreements were settled through consultation with an experienced reviewer (RJJ).

| Statistical analysis
Among the included studies, different measurement tools were used to evaluate the pruritus scores and quality of life; thus, the standardized mean difference (SMD) was calculated.The SCORAD scores and frequency of TCS were evaluated using the same tools, therefore mean difference (MD) was used.The risk difference (RD) was calculated due to no adverse events.The uncertainty was expressed with 95% confidence intervals (CIs).Heterogeneity was measured using the chi-squared test and I 2 statistic. 25

| Subgroup analysis
Subgroup analyses were conducted according to the types of MA 26 (a single strain of probiotic, the mixture of probiotic strains, prebiotics or synbiotics), the age of participants 27 (infants, children or adults) and the severity of AD 28 (mild to moderate, moderate or moderate to severe).

| Meta regression analysis
Univariable meta-regression analyses were used to investigate potential sources of heterogeneity based on the characteristics of the study (MA types, participants' age, and AD severity).

| Sensitivity analysis
We performed sensitivity analyses by eliminating studies one by one to verify the robustness of the pooled results.

| Publication bias
We used a funnel plot and Egger's test to detect publication bias when ≥10 studies with the same outcome were included in the analysis.

| TSA
We assessed the risk of false positives or false negatives for primary outcomes by TSA (version 0.9.5.10-Beta) in a meta-analysis. 29Random effects model with a maximum type I error of 5%, and a maximum type II error of 20% (80% power) were applied.When the cumulative Zcurve enters the useless zone or crosses the trial sequential monitoring boundary, the expected intervention effect may reach an adequate level of evidence.If the Z-curve does not cross any XUE ET AL. boundaries and does not reach the required size of information, it indicates that the evidence is insufficient to draw a conclusion.

| Certainty of evidence
We applied the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) 30 tool to assess the certainty of evidence.Each outcome was evaluated based on the following five aspects: limitations, inconsistency, indirectness, imprecision, and publication bias.The certainty of evidence was accordingly graded as "high," "moderate," "low," or "very low".

| Risk of bias assessment
The plot of the risk of bias (RoB 2.0) for each included study is presented in Figure 2A, and the proportions of individual studies are presented in Figure 2B.In the randomization process, deviation from intended interventions, measurement of outcomes and selection of the reported result, all trials were rated as low risk of bias.For missing outcome, six studies 14,16,36,41,44,45 with high dropout rates and did not report the details of drop-outs, which were rated as high risk of bias.In summary, the overall risk of bias in 15 trials was considered low risk and six trials were considered high.

| Results of subgroup analysis
We conducted subgroup analyses according to different MA types (Figure S1 and S2 in Appendix 3), participants' age (Figure S3 and S4 in Appendix 3) and AD severity (Figure S5 and S6 in Appendix 3).

| Subgroup of different MA types
In the outcome of SCORAD scores (Table 2), oral mixtures of probiotic strains and prebiotics were more effective than TCS.
Furthermore, oral mixtures of probiotic strains were better than other MA types.In terms of adverse events (Table 3), all types of the MA were not superior to TCS, except synbiotic.  - of 17

| Subgroup of different participants' age
Oral MA reduced the SCORAD scores (Table 2) in children but had no effect on infants.With regard to adverse events (Table 3), there were no statistical differences between MA þ TCS and TCS in AD patients at different ages.

| Subgroup of different AD severity
The results of subgroup analysis demonstrated that oral MA plus TCS had an effect on decreasing SCORAD scores in moderate and moderate to severe AD patients (Table 2).However, there was no effect on adverse events among AD patients with different severity (Table 3).

| Sensitivity analysis
The results of SCORAD scores and adverse events did not alter after excluding studies one by one (Figures 7-8 in Appendix 3).

| Results of meta regression analysis
The results of univariate meta-regression showed that the types of MA and the severity of AD were the sources of heterogeneity (Table 4).

| Publication bias
The funnel plot and Egger's test of SCORAD scores and adverse events suggested that no evidence of publication bias existed (Figuress 8A-B).

| TSA
Although the cumulative sample size did not meet expectations, the Zcurve of SCORAD crossed both the conventional and TSA boundaries, which indicated that MA, as an add-on therapy to TCS, was superior to TCS (Figure S9, Appendix 3).The included sample size of adverse events did not achieve the required information size, and its cumulative Z-curve did not cross either the conventional boundaries or the

| Certainty of evidence
The results of certainty of evidence are shown in Figure S11, Appendix 3. The adverse events were rated as high certainty of evidence.The certainty of evidence for pruritus scores, quality of life and the frequency of TCS were graded as moderate, while the evidence of the SCORAD was rated as low certainty.The reasons for downgrading were mainly attributed to the risk of bias of the included studies, inconsistency and imprecision.

| DISCUSSION
In the present study, the results showed that MA as an add-on therapy was better than TCS in lowering the SCORAD scores.
However, the high heterogeneity (I 2 = 81%) was detected, which -13 of 17 should be treated with caution.The results of meta-regression indicated that different MA types and AD severity were the reasons for heterogeneity.The minimum clinically important difference (MCID) is the smallest change in the outcome measure and refers to a clinically relevant outcome. 48Researchers 49,50 reported that the reduction of the SCORAD index to more than 8 units was considered as MCID.The pooled result of SCORAD scores in this study was a reduction of 5 units.It might be associated with the inconsistency of disease severity and probiotic strains. 51According to the results of subgroup analysis on MA types, oral mixed strains and synbiotic decreased the SCORAD index by 9.35 and 16.3 units, respectively.With regard to AD severity, the SCORAD index in patients with moderate AD decreased 12.09 units.The results of adverse events indicated that MA plus TCS was a safe way to treat AD.There were no differences in pruritus, quality of life, and frequency of TCS using in comparison of MA plus TCS versus TCS.
It was reported that immune regulation and anti-inflammatory effects of MA played an important role in the treatment of AD. 52 Rosenfeldt et al 36 observed that serum eosinophil cationic protein levels decreased in AD patients who were administered MA orally.
Serum eosinophil cationic protein, a cytotoxic protein released by activated eosinophils, rises rapidly during acute exacerbations of AD and is considered to be an indicator of AD at acute exacerbations.Prakoeswa et al 35 found that patients treated with MA plus TCS had lower levels of interleukin-4 and interferon gamma and higher levels of interleukin-10.Kim et al 53 discovered that oral MA could reduce AD-associated skin lesions, epidermal thickening, serum levels of immunoglobulin E, and immune cell infiltration.Therefore, we speculated that the therapeutic-boosting effect of MA may be associated with the modulation of inflammatory factors and immunoreactive molecules.However, the specific mechanism of MA for AD is still unclear and needs to be further studied.

| Implications for clinical practice
With regards to the types of MA, a mixture of probiotics (Lactobacillus and Bifidobacterium combined) was more effective than probiotics alone, which achieved the MCID in SCORAD index.Jiang et al. 54 and Uwaezuoke et al. 55 discovered that probiotics could relieve AD symptoms, especially using mixed-strain probiotics (Lactobacillus and Bifidobacterium).Another meta-analysis 56 also favored that mixed strains of Lactobacillus and Bifidobacterium could reduce the incidence of AD.Thus, a mixed-strain probiotic component might be beneficial to patients with AD.The duration of MA ranged from 6 to 24 weeks, most of which were administered MA orally for 8 weeks or 12 weeks.Dosage form included tablets, liquids, powders, and capsules.The majority of the included studies reported that oral MA doses were 5-10 � 10 9 colony-forming units/day (CFU/day).CFU/ day, a measure of the MA dose, was related to the positive effects if there were more than 10 8 CFU/day. 57However, due to limited available studies, the clinical application of MA is not fully understood yet. 31,42Future research should focus on the optimal clinical protocols of MA treatment for AD.
In addition, the results of subgroup analyses showed that MA ameliorated the symptoms of AD in children, but not in infants and adults.In contrast to adults, the gut microbiota of children was relatively unstable, which was vulnerable to external factors such as nutrition, diet and external environments. 58,59Infants had a monodiet and their gut microbiota was mainly derived from breastfeeding or formula. 60Different feeding practices might affect the fixation of oral MA in the infant gut.Besides, oral MA supplementation had a therapeutic-boosting effect in moderate or severe AD patients receiving TCS treatment.In patients with mild AD, the effect of MA could be masked by potent TCS.TCS can control the symptoms of most mild to moderate AD patients, but not severe AD patients. 6MA as a complementary therapy to TCS treatment might be a promising treatment option for patients with moderate or severe AD.We were also concerned about whether MA, as an adjunct therapy, had a steroid sparing effect.It would be beneficial for reducing the corticophobia and improving compliance with the therapeutic regimen.In the present study, no difference was detected in the frequency and grams of TCS after in combination with MA.
Since few studies 42,43 focused on it, the result should be treated with caution.Meanwhile, due to limited studies, the steroid sparing effect in different ages is still unclear.More studies are needed to address this issue.

| LIMITATIONS
Several potential limitations of the present meta-analysis should be acknowledged.First, the number of included RCTs was limited, and the sample size of each study was small.Second, the relevant articles published in English were retrieved, and selection bias was inevitable.
Third, owing to limited studies, the optimal protocols of MA plus TCS therapy were not determined (the optimal types of MA, the optimal dosage, frequency, et al.).

| CONCLUSION
Oral MA plus TCS could be an effective and safe treatment for patients with AD to relieve relevant symptoms, which might be used as an add-on therapy in the treatment of AD.Owing to limited available studies, results should be interpreted with caution.Further studies with a larger sample size are needed to explore the optimal protocol of MA plus TCS.

( 1 )
study information: first author, publication year, and country; (2) participant characteristics: severity, sample size, and age; (3) details of interventions and comparators: types, regimens (dose and dosage form) and duration; (4) primary outcomes and secondary outcomes; and (6) Main results.Any discrepancies were arbitrated by a third reviewer (JL).

F I G U R E 2
(A) The plot of RoB 2.0 for each included study.(B) Proportions of individual study for each domain.

3 F I G U R E 4 F I G U R E 5
Forest plot of SCORSD scores.Forest plot of adverse events.Forest plot of pruritus scores.F I G U R E 6 Forest plot of quality of life.XUE ET AL.

F I G U R E 8
(A) Funnel plot of SCORSD scores.(B) Funnel plot of adverse events.
Characteristics of the included studies.
Flow chart for selection of eligible studies.T A B L E 1 a mixture of probiotics (Lactobacillus rhamnosus, Lactobacillus acidophilus, Lactobacillus paracasei, and Bifidobacterium lactis.However, this clinical benefit is related to treatment duration.Probiotics should be considered as an adjuvant treatment for AD.-XUE ET AL.T A B L E 1 (Continued)T A B L E 1 (Continued) E 7 Forest plot of the frequency of TCS.Subgroup analysis of SCORAD.
Subgroup Analysis of Adverse events.Results of univariable meta-regression.
T A B L E 3